Detection of Bean Pod Mottle Virus RT-PCR Amplicon by Dipstick Assay(PDF)

《大豆科学》[ISSN:1000-9841/CN:23-1227/S]

Issue:

2010年06期

Page:

1024-1027

Research Field:

Publishing date:

Info

Title:

Detection of Bean Pod Mottle Virus RT-PCR Amplicon by Dipstick Assay

Author(s):

CAO Cheng¹;  WEI Mei-sheng²;  WU Xing-quan¹;  ZHANG Yong-jiang²;  LI Gui-fen²

1.College of Bioengineering, Henan University of Technology, Zhengzhou 450001，Henan;

2.Institute of Animal and Plant Quarantine, Chinese Academy of Inspection and Quarantine, Beijing 100029, China

Keywords:

Colloidal gold; Dipstick; BPMV; RT-PCR ; Nucleic acid detection

PACS:

S565.1

DOI:

10.11861/j.issn.1000-9841.2010.06.1024

Abstract:

Bean pod mottle virus (BPMV) is a quarantine pest for China. In order to detect the virus on site, we have developed a rapid method for detection of RT-PCR amplicon of the virus. The method is performed by labeling one prime with biotin the other primer with fluorescein (or digoxigenin). The dual-labeled amplicon can be generated from routine polymerase chain reaction. Rabbit polyclonal antibody against biotin is conjugated with 20~30 nm colloidal gold particles fixed in a conjugate pad.Monoclonal antibody against fluorescein (or digoxigenin) is spotted on nitrocellulose membrane as T dot. Goat anti-rabbit polyclonal antibody is spotted on nitrocellulose membrane as C dot. The RT-PCR amplicon is detected on the test dot(T dot), while the C dot serves as a control. Using this method we have detected the RT-PCR amplicon of bean pod mottle virus within 15 minutes without the staining of ethidium bromide and the running of agarose gel.

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